Pulmonary vasculature is anatomically predisposed to deposition of fibrin and thromboemboli formed in the vasculature (for example, upon deep vein thrombosis). Both emboli and fibrin lodged in the lung play an important role in the pulmonary and cardiovascular pathology and contribute significantly to morbidity and mortality of disease conditions including, but not limited to, thrombosis, atherosclerosis, deep vein thrombosis, diabetes, adult respiratory distress syndrome, pulmonary embolism, shock and sepsis. Anticoagulants (e.g., heparin) are useful in preventing formation of intravascular fibrin clots, whereas fibrinolytics (e.g., plasminogen activators) are useful for dissolution of fibrin clots. Both anticoagulants and fibrinolytics, however, undergo inactivation and elimination from the bloodstream. This restricts their applicability for treatment of pulmonary embolism. Administration of large doses and/or multiple injections of a drug to compensate for elimination/inactivation impose inconvenience in treatment and high risk of harmful side effects. Uncontrolled bleeding is an example of such side effects of prolonged administration or a large dose of anticoagulants or fibrinolytics.
Augmentation of anticoagulant or/and fibrinolytic potential of the luminal surface of endothelial cells lining pulmonary vessels thus represents an important therapeutic strategy for treatment or/and prevention of disease conditions associated with or manifested by pulmonary embolism and fibrin deposition. Because these therapeutics must have access to the blood components in order to control coagulation or activate fibrinolysis, a requirement for such a strategy is that the anticoagulant or fibrinolytic agent be associated for a prolonged time with the luminal surface of the pulmonary endothelium.
One approach to attain this objective is to conjugate a drug to an antibody against surface endothelial molecules. This conjugation provides selective delivery, also referred to herein as targeting, of a drug to endothelium and prolonged association of a drug with endothelium. Therapeutic enzymes and genetic material conjugated to such antibodies have been demonstrated to bind to the endothelial cells in vitro and in vivo after injection in animals. Since the lungs contain approximately 30% of the total amount of endothelial cells in the body and receive a whole cardiac output of venous blood, antibodies against endothelial antigens tend to accumulate in the lung after intravenous injection. For example, Kennel et al. have described an antibody against thrombomodulin which recognizes endothelial surface in vivo, accumulates in the pulmonary vasculature and is capable of delivery of conjugated liposomes to the pulmonary endothelium (Kennel et al. 1990 Nucl. Med. Biol. 17:193-100; Trubetskoy et al. 1992 Biochim. Biophys. Acta 1131:311-313). An antibody against angiotensin-converting enzyme (ACE) has been described which possesses very similar properties (Danilov et al. 1991 Lab. Invest. 64:118-124). Therapeutic enzymes such as catalase, superoxide dismutase and plasminogen activators conjugated with ACE antibody have been demonstrated to accumulate in the lungs after intravascular injection (Muzykantov et al. 1996 Proc. Nat'l Acad. Sci. USA 93:5213-5218; Muzykantov et al. 1997 J. Pharm. Exp. Therap. 279:1026-1034). In addition, an antibody against E-selectin has been described which binds to and delivers liposomes to the cytokine-activated endothelium in cell culture (Spragg et al. 1997 Proc. Nat'l Acad. Sci. USA 94:8795-8800). A PECAM antibody conjugated with streptavidin has also been recently described which provides an effective carrier for delivery of drugs to the endothelium (Muzykantov et al. 1998 Am. J. Resp. Crit. Care Med. 157:A203).
However, endothelial cells internalize antibodies against thrombomodulin (Muzykantov et al. 1997 Circulation 96:I43-44), ACE (Muzykantov et al. 1996 Am. J. Physiol. 270:L704-713), E-selectin (Spragg et al. 1997 Proc. Nat'l Acad. Sci. USA 94:8795-8800) and anti-PECAM/streptavidin complex (Muzykantov et al. 1998 Am. J. Resp. Crit. Care Medicine 157:A203). Thus, while these carriers provide intracellular delivery, a feature which may be useful for targeting of genes and some other therapeutic agents, anticoagulants or fibrinolytics must escape internalization and remain on the luminal surface in order to control blood components. Accordingly, these carrier antibodies are of limited use in the delivery of anticoagulants, fibrinolytics or other drugs wherein their therapeutic action is localized to the blood.
An ICAM-1 monoclonal antibody, mAb 1A29 has also been described which accumulates in rat lungs following i.v. injection. Conjugation of catalase to this ICAM-1 monoclonal antibody via a streptavidin-biotin crosslinker resulted in accumulation of the anti-ICAM-1 conjugated catalase in the lung and protection of the lung from damage by extracellular oxidants (Muzykantov et al. Am. J. Resp. Crit. Care Medicine 1997 155(4):p. A187). Radiolabeled mAb 1A29 has also been shown to accumulate in the vasculature challenged with pro-inflammatory agents TNF and endotoxin (Mulligan et al. 1993 Am. J. Pathol. 142:1739-1749). In addition, this antibody has been shown to react with normal endothelial cells in the rat vasculature and that injection of TNF or endotoxin stimulates endothelial binding of mAb 1A29 (Panes et al. 1995 Am. J. Physiol. 269:H1955-1964). This antibody has also been shown to attenuate vascular injury induced by activated leukocytes via blocking of their adhesion to the endothelial cells.
It has now been found that monoclonal antibodies against the endothelial surface antigen ICAM-1 bind effectively to the endothelial cells without subsequent internalization. Conjugation of a drug to an non-internalizable antibody such as the ICAM-1 monoclonal antibody which binds to an antigen on the luminal surface of the pulmonary vasculature provides a useful means for targeted delivery and retention of the drug on the luminal surface, or blood compartment, of the pulmonary vasculature.